Glutamate receptor-mediated calcium entry in neurons derived from P19 embryonal carcinoma cells

L. M.T. Canzoniero, S. L. Sensi, D. M. Turetsky, M. F.A. Finley, D. W. Choi, James E. Huettner

Research output: Contribution to journalArticle

15 Scopus citations

Abstract

We have examined the control of calcium elevation by glutamate in neurons derived from the mouse P19 embryonal carcinoma cell line. Following transient exposure to retinoic acid, P19 cells differentiate into neurons that express both NMDA and non-NMDA glutamate receptor subtypes. Fluorescence videomicroscopy using the indicator fura-2 revealed concentration-dependent elevation in cytosolic calcium levels with exposure to NMDA or kainate. Replacement of extracellular sodium with N-methylglucamine significantly reduced the action of kainate. Exposure to high K + medium also elicited an elevation of cytosolic calcium in P19 cells, which was partially inhibited by the calcium channel antagonist nimodipine. These experiments suggest that the elevation in calcium produced by kainate involves the activation of voltage- gated calcium channels as a consequence of membrane depolarization, in contrast to direct calcium entry through NMDA receptor channels. Whole-cell recordings revealed that P19 NMDA receptors were highly permeable to calcium (P(Ca)/P(Na) = 5.6 ± 0.2). In most cells, channels gated by kainate displayed low permeability to calcium; the median permeability ratio, P(Ca)/P(Na), was 0.053 (range 0.045 to 0.132). Activation of peak currents by NMDA, glycine, and kainate was half-maximal at 24 μM, 240 nM, and 81 μM, respectively. In addition, cadmium-sensitive currents through voltage-gated calcium channels were recorded in P19 cells bathed in barium/TEA chloride. Staining with antibodies directed against AMPA receptor subunits revealed widespread immunoreactivity for anti-GluR-B/C and anti-GluR-B/D. About half of the P19 cells were stained with antibodies selective for GluR-D but there was little or no immunoreactivity for the GluR-A subunit.

Original languageEnglish
Pages (from-to)226-236
Number of pages11
JournalJournal of Neuroscience Research
Volume45
Issue number3
DOIs
StatePublished - Oct 7 1996

Keywords

  • N-methyl-D-aspartate
  • excitotoxicity
  • fura-2
  • mouse
  • permeation

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